Keratinocyte derived extracellular vesicles mediated crosstalk between epidermis and dermis in UVB-induced skin inflammation.

BACKGROUND AND RATIONALE: Ultraviolet-B (UVB) light induces dermal inflammation, although it is mostly absorbed in the epidermis. Recent reports suggest extracellular vesicles (EVs) act as a mediator of photodamage signaling. Melatonin is reported to be a protective factor against UV-induced damage. We hypothesized that EVs derived from UVB-irradiated keratinocytes might trigger proinflammatory responses in dermal cells and tested whether melatonin can ameliorate UVB-induced inflammation. METHODS: We used UVB-irradiated HaCaT cells, primary keratinocytes and STING knock-out mice to model production of EVs under photodamaging conditions and performed immunoblotting and ELISA to measure their effect on dermal macrophages. RESULTS: UVB-irradiated keratinocytes produce an increased number of EVs that contain higher concentrations of DNA and protein compared with controls. KC-derived EVs (KEVs) induced a STING- and inflammasome-mediated proinflammatory response in macrophages in vitro, and a pronounced inflammatory infiltrate in mouse dermis in vivo. Melatonin ameliorated KEVs inflammatory effect both in vitro and in vivo. CONCLUSIONS: This data suggests EVs are mediators in a crosstalk that takes place between keratinocytes and their neighboring cells as a result of photodamage. Further studies exploring EVs induced by damaging doses of UVB, and their impact on other cells will provide insight into photodamage and may help develop targeted therapeutic approaches.

Two novel anti-aminoacyl tRNA synthetase antibodies: Autoantibodies against cysteinyl-tRNA synthetase and valyl-tRNA synthetase.

Anti-aminoacyl-tRNA synthetase (anti-ARS) antibodies are useful for identifying a clinical subset of patients with inflammatory myopathies. Since the myositis of anti-ARS-positive patients is characterized by a unique set of non-myopathic manifestations, including interstitial lung disease, mechanic's hands, and arthralgia, the patients are classified as having anti-synthetase syndrome. Autoantibodies have been identified to eight kinds of ARSs. Of the other 12 ARSs, eight are components of the "OJ" multi-synthetase complex. Autoantibodies to the four remaining ARSs (CysARS, ValARS, SerARS, and TrpARS) have not been reported to be present in patients with inflammatory myopathies. In this study, we first screened samples from more than 300 Japanese patients majorly consisting of those with dermatomyositis (DM) by our established in-house ELISA to find autoantibodies against the four ARSs described above. Since sera from two DM patients specifically reacted to CysARS or ValARS, we determined their reactivities by immunoprecipitation (IP) with the corresponding recombinant proteins and IP-Western blotting with cellular extract. One patient had several features found in anti-synthetase syndrome, but the other did not. The clinical differences among the various anti-ARS antibodies should be explored in a future work.

Clinical significance of anti-NOR90 antibodies in systemic sclerosis and idiopathic interstitial pneumonia.

OBJECTIVE: Anti-NOR90 antibodies are usually found in patients with SSc; however, their clinical relevance remains obscure. We developed an ELISA for measuring them to investigate the clinical features of patients with anti-NOR90 antibodies. METHODS: Serum samples from 1252 patients with various conditions from Nagoya University Hospital and 244 patients with idiopathic interstitial pneumonia (IIP) from Tosei General Hospital were included. Anti-NOR90 antibodies were assayed by an ELISA using the recombinant protein produced by in vitro transcription/translation. RESULTS: Five (0.4%) patients in the Nagoya University Hospital cohort had anti-NOR90 antibodies. One patient with diffuse cutaneous SSc, three with limited cutaneous SSc, and one with Raynaud's disease were positive for anti-NOR90 antibodies. Anti-NOR90 antibodies were found more frequently in patients with systemic scleroderma-spectrum disorders (SSDs) than without SSDs (5/316 vs 0/936, P <0.00101) and were found more frequently in patients with SSc than without SSc (4/249 vs 0/528, P <0.0104) in the systemic autoimmune rheumatic diseases cohort. Three of the four anti-NOR90-positive SSc patients had interstitial lung disease (ILD), and two of those four had cancer. Three (1.2%) patients in the Tosei General Hospital cohort had anti-NOR90 antibodies. All three of the anti-NOR90-positive IIP patients had gastrointestinal tract involvement, and two of those three had cancer or skin lesions observed in SSc. CONCLUSIONS: Although anti-NOR90 antibodies are rarely found in clinics, our ELISA is useful for their detection. Further studies are needed to confirm the association of anti-NOR90 antibodies with ILD and cancer in SSc and IIP patients.

Strong correlation between cancer progression and anti-transcription intermediary factor 1γ antibodies in dermatomyositis patients.

OBJECTIVES: Transcription intermediary factor 1γ (ΤΙF1γ) protein is known as a tumour suppressor that promotes cellular differentiation. Autoantibodies to ΤΙF1γ have a strong clinical association with cancers associated with dermatomyositis (DM). This study aims to identify the clinical characteristics of cancers in anti-ΤΙF1γ antibody-positive adult patients with DM. METHODS: This retrospective analysis covered 160 adult DM patients who visited Nagoya University Hospital or collaborating medical centres between 2003 and 2016. Anti-TIF1γ antibody and other myositis-specific autoantibodies were detected by ELISA. Based on a review of medical charts, the cancers were staged according to the TNM Classification of Malignant Tumours of the Union for International Cancer Control and were divided into the two groups of "advanced" or "non-advanced" according to the stage classification. RESULTS: Forty-one of the 160 (26%) patients had cancer. The incidence was significantly higher in the anti-TIF1γ-positive patients than in the anti-TIF1γ-negative patients (23/34=68% vs. 18/126=14%, p<1x10-6). Anti-TIF1γ-positive patients with cancer were found more frequently in the "advanced" group than in the "non-advanced" group (21/23=91% vs. 9/18=50%, p<0.0046). The intervals between DM diagnosis and cancer diagnosis were significantly shorter in the anti-TIF1γ-positive patients than in the anti-TIF1γ-negative patients (p=0.047). CONCLUSIONS: Not only did anti-TIF1γ antibodies correlate strongly with malignancy in DM patients, but cancers were also significantly more advanced in anti-TIF1γ-positive DM patients than in anti-TIF1γ-negative patients. Cancers in such cases were very frequently found close to the time of the DM diagnosis.

Splicing variant of WDFY4 augments MDA5 signalling and the risk of clinically amyopathic dermatomyositis.

OBJECTIVES: Idiopathic inflammatory myopathies (IIMs) are a heterogeneous group of rare autoimmune diseases in which both genetic and environmental factors play important roles. To identify genetic factors of IIM including polymyositis, dermatomyositis (DM) and clinically amyopathic DM (CADM), we performed the first genome-wide association study for IIM in an Asian population. METHODS: We genotyped and tested 496 819 single nucleotide polymorphism for association using 576 patients with IIM and 6270 control subjects. We also examined the causal mechanism of disease-associated variants by in silico analyses using publicly available data sets as well as by in in vitro analyses using reporter assays and apoptosis assays. RESULTS: We identified a variant in WDFY4 that was significantly associated with CADM (rs7919656; OR=3.87; P=1.5×10-8). This variant had a cis-splicing quantitative trait locus (QTL) effect for a truncated WDFY4isoform (tr-WDFY4), with higher expression in the risk allele. Transexpression QTL analysis of this variant showed a positive correlation with the expression of NF-κB associated genes. Furthermore, we demonstrated that both WDFY4 and tr-WDFY4 interacted with pattern recognition receptors such as TLR3, TLR4, TLR9 and MDA5 and augmented the NF-κB activation by these receptors. WDFY4 isoforms also enhanced MDA5-induced apoptosis to a greater extent in the tr-WDFY4-transfected cells. CONCLUSIONS: As CADM is characterised by the appearance of anti-MDA5 autoantibodies and severe lung inflammation, the WDFY4 variant may play a critical role in the pathogenesis of CADM.